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International Journal of Molecular... Jul 2022Bacterial cellulose is a natural polymer with an expanding array of applications. Because of this, the main cellulose producers of the genus have been extensively...
Bacterial cellulose is a natural polymer with an expanding array of applications. Because of this, the main cellulose producers of the genus have been extensively studied with the aim to increase its synthesis or to customize its physicochemical features. Up to now, the genetic studies in have focused on the first cellulose synthase operon () encoding the main enzyme complex. However, the role of other accessory cellulose operons has been understudied. Here we aimed to fill this gap by performing a detailed analysis of the second cellulose synthase operon (), which is putatively linked with cellulose acylation. In this study we harnessed the genome sequence, gene expression and protein structure information of E25 and other species to discuss the probable features of and the biochemical function of its main protein products. The results of our study support the previous hypothesis that is involved in the synthesis of the acylated polymer and expand it by presenting the evidence that it may also function in the regulation of its attachment to the cell surface and to the crystalline cellulose fibers.
Topics: Acetobacteraceae; Cellulose; Gluconacetobacter xylinus; Glucosyltransferases; Operon
PubMed: 35887199
DOI: 10.3390/ijms23147851 -
Applied Microbiology Jan 1964The interaction between photosynthetic microorganisms and an inert electrode material was examined. Cathodic polarization values of platinum-bearing marine algae were...
The interaction between photosynthetic microorganisms and an inert electrode material was examined. Cathodic polarization values of platinum-bearing marine algae were obtained over a wide current-density range under both illumination and dark conditions. A potential shift of 0.6 v in the cathodic direction occurred upon illumination at a current density of 4.3 mua/cm(2). Similar photo-induced results, involving anodic polarization, were obtained by use of resting cells of Rhodospirillum rubrum supplemented with malate. Appropriate combinations of such bioelectrodes were used to assemble an electrochemical cell capable of light-dependent production of electrical energy.
Topics: Electrophysiology; Eukaryota; Light; Malates; Metabolism; Photosynthesis; Platinum; Research; Rhodospirillum; Rhodospirillum rubrum
PubMed: 14106931
DOI: 10.1128/am.12.1.10-12.1964 -
Journal of Bacteriology Jan 2021Cellulose is a widespread component of bacterial biofilms, where its properties of exceptional water retention, high tensile strength, and stiffness prevent dehydration...
Cellulose is a widespread component of bacterial biofilms, where its properties of exceptional water retention, high tensile strength, and stiffness prevent dehydration and mechanical disruption of the biofilm. Bacteria in the genus secrete crystalline cellulose, with a structure very similar to that found in plant cell walls. How this higher-order structure is produced is poorly understood. We used cryo-electron tomography and focused-ion-beam milling of native bacterial biofilms to image cellulose-synthesizing and bacteria in a frozen-hydrated, near-native state. We confirm previous results suggesting that cellulose crystallization occurs serially following its secretion along one side of the cell, leading to a cellulose ribbon that can reach several micrometers in length and combine with ribbons from other cells to form a robust biofilm matrix. We were able to take direct measurements in a near-native state of the cellulose sheets. Our results also reveal a novel cytoskeletal structure, which we have named the cortical belt, adjacent to the inner membrane and underlying the sites where cellulose is seen emerging from the cell. We found that this structure is not present in other cellulose-synthesizing bacterial species, and 1094, which do not produce organized cellulose ribbons. We therefore propose that the cortical belt holds the cellulose synthase complexes in a line to form higher-order cellulose structures, such as sheets and ribbons. This work's relevance for the microbiology community is twofold. It delivers for the first time high-resolution near-native snapshots of spp. (previously spp.) in the process of cellulose ribbon synthesis, in their native biofilm environment. It puts forward a noncharacterized cytoskeleton element associated with the side of the cell where the cellulose synthesis occurs. This represents a step forward in the understanding of the cell-guided process of crystalline cellulose synthesis, studied specifically in the genus and still not fully understood. Additionally, our successful attempt to use cryo-focused-ion-beam milling through biofilms to image the cells in their native environment will drive the community to use this tool for the morphological characterization of other studied biofilms.
Topics: Acetobacteraceae; Biofilms; Cellulose; Crystallization; Cytoskeleton; Electron Microscope Tomography; Electrons; Escherichia coli; Gluconacetobacter; Gluconacetobacter xylinus; Microfibrils
PubMed: 33199282
DOI: 10.1128/JB.00371-20 -
Microbial Cell Factories Feb 2021Because of its tractability and straightforward cultivation, the magnetic bacterium Magnetospirillum gryphiswaldense has emerged as a model for the analysis of...
BACKGROUND
Because of its tractability and straightforward cultivation, the magnetic bacterium Magnetospirillum gryphiswaldense has emerged as a model for the analysis of magnetosome biosynthesis and bioproduction. However, its future use as platform for synthetic biology and biotechnology will require methods for large-scale genome editing and streamlining.
RESULTS
We established an approach for combinatory genome reduction and generated a library of strains in which up to 16 regions including large gene clusters, mobile genetic elements and phage-related genes were sequentially removed, equivalent to ~ 227.6 kb and nearly 5.5% of the genome. Finally, the fragmented genomic magnetosome island was replaced by a compact cassette comprising all key magnetosome biosynthetic gene clusters. The prospective 'chassis' revealed wild type-like cell growth and magnetosome biosynthesis under optimal conditions, as well as slightly improved resilience and increased genetic stability.
CONCLUSION
We provide first proof-of-principle for the feasibility of multiple genome reduction and large-scale engineering of magnetotactic bacteria. The library of deletions will be valuable for turning M. gryphiswaldense into a microbial cell factory for synthetic biology and production of magnetic nanoparticles.
Topics: Gene Deletion; Genome, Bacterial; Magnetosomes; Magnetospirillum
PubMed: 33541381
DOI: 10.1186/s12934-021-01517-2 -
Research in Microbiology 2022Cadmium (Cd) is a heavy metal used as raw material for several fertilizers and pesticides. The increase of Cd concentration in soils has been observed in cultivated...
Cadmium (Cd) is a heavy metal used as raw material for several fertilizers and pesticides. The increase of Cd concentration in soils has been observed in cultivated areas, affecting animals, plants, and microorganisms. Gluconacetobacter diazotrophicus is a plant growth-promoting bacterium able to survive under adverse environmental conditions. Here, we investigated key mechanisms involved with the resistance of G. diazotrophicus to Cd. Proteomic analyses revealed that the main pathways regulated in response to Cd are nutrient uptake, multidrug efflux pumps, response to oxidative stress, and protein quality control system. Extracytoplasmic proteins related to multidrug efflux pumps were up-accumulated, while several proteins related to nutrients uptake were down-accumulated. The relevance of these pathways for bacterial resistance to Cd was investigated by reverse genetic analysis using mutants defective for nutrient uptake (tdbr, ompW, and oprB), multidrug efflux (czcC), response to oxidative stress (ggt), and protein quality control system (clpX). Our data demonstrated the essential role of the tdbr and czcC genes for resistance to Cd in G. diazotrophicus. These results contribute to a better understanding of the resistance mechanisms to Cd in G. diazotrophicus, shedding light on responses associated with extracytoplasmic compartments.
Topics: Cadmium; Gluconacetobacter; Plants; Proteomics
PubMed: 35104604
DOI: 10.1016/j.resmic.2022.103922 -
Applied and Environmental Microbiology Sep 2020Purple nonsulfur bacteria are increasingly recognized for industrial applications in bioplastics, pigment, and biomass production. In order to optimize the yield of...
Purple nonsulfur bacteria are increasingly recognized for industrial applications in bioplastics, pigment, and biomass production. In order to optimize the yield of future biotechnological processes, the assimilation of different carbon sources by has to be understood. As they are released from several fermentation processes, volatile fatty acids (VFAs) represent a promising carbon source in the development of circular industrial applications. To obtain an exhaustive characterization of the photoheterotrophic metabolism of in the presence of valerate, we combined phenotypic, proteomic, and genomic approaches. We obtained evidence that valerate is cleaved into acetyl coenzyme A (acetyl-CoA) and propionyl-CoA and depends on the presence of bicarbonate ions. Genomic and enzyme inhibition data showed that a functional methylmalonyl-CoA pathway is essential. Our proteomic data showed that the photoheterotrophic assimilation of valerate induces an intracellular redox stress which is accompanied by an increased abundance of phasins (the main proteins present in polyhydroxyalkanoate [PHA] granules). Finally, we observed a significant increase in the production of the copolymer P(HB--HV), accounting for a very high (>80%) percentage of HV monomer. Moreover, an increase in the PHA content was obtained when bicarbonate ions were progressively added to the medium. The experimental conditions used in this study suggest that the redox imbalance is responsible for PHA production. These findings also reinforce the idea that purple nonsulfur bacteria are suitable for PHA production through a strategy other than the well-known feast-and-famine process. The use and the littering of plastics represent major issues that humanity has to face. Polyhydroxyalkanoates (PHAs) are good candidates for the replacement of oil-based plastics, as they exhibit comparable physicochemical properties but are biobased and biodegradable. However, the current industrial production of PHAs is curbed by the production costs, which are mainly linked to the carbon source. Volatile fatty acids issued from the fermentation processes constitute interesting carbon sources, since they are inexpensive and readily available. Among them, valerate is gaining interest regarding the ability of many bacteria to produce a copolymer of PHAs. Here, we describe the photoheterotrophic assimilation of valerate by , a purple nonsulfur bacterium mainly known for its metabolic versatility. Using a knowledge-based optimization process, we present a new strategy for the improvement of PHA production, paving the way for the use of in industrial processes.
Topics: Heterotrophic Processes; Phototrophic Processes; Polyhydroxyalkanoates; Rhodospirillum rubrum; Valerates
PubMed: 32651203
DOI: 10.1128/AEM.00901-20 -
Bacteriological Reviews Sep 1977
Review
Topics: Bacteria; Cell Cycle; Cell Division; Models, Biological; Morphogenesis; Rhodospirillaceae; Species Specificity
PubMed: 334156
DOI: 10.1128/br.41.3.754-808.1977 -
Nature Communications Aug 2021Engineered living materials (ELMs) based on bacterial cellulose (BC) offer a promising avenue for cheap-to-produce materials that can be programmed with genetically...
Engineered living materials (ELMs) based on bacterial cellulose (BC) offer a promising avenue for cheap-to-produce materials that can be programmed with genetically encoded functionalities. Here we explore how ELMs can be fabricated in a modular fashion from millimetre-scale biofilm spheroids grown from shaking cultures of Komagataeibacter rhaeticus. Here we define a reproducible protocol to produce BC spheroids with the high yield bacterial cellulose producer K. rhaeticus and demonstrate for the first time their potential for their use as building blocks to grow ELMs in 3D shapes. Using genetically engineered K. rhaeticus, we produce functionalized BC spheroids and use these to make and grow patterned BC-based ELMs that signal within a material and can sense and report on chemical inputs. We also investigate the use of BC spheroids as a method to regenerate damaged BC materials and as a way to fuse together smaller material sections of cellulose and synthetic materials into a larger piece. This work improves our understanding of BC spheroid formation and showcases their great potential for fabricating, patterning and repairing ELMs based on the promising biomaterial of bacterial cellulose.
Topics: Acetobacteraceae; Bioengineering; Biofilms; Cellulose; Genetic Engineering; Regenerative Medicine
PubMed: 34413311
DOI: 10.1038/s41467-021-25350-8 -
Applied and Environmental Microbiology Apr 2018Few data have been published on the occurrence and functional role of acetic acid bacteria (AAB) in lambic beer production processes, mainly due to their difficult...
Temporal and Spatial Distribution of the Acetic Acid Bacterium Communities throughout the Wooden Casks Used for the Fermentation and Maturation of Lambic Beer Underlines Their Functional Role.
Few data have been published on the occurrence and functional role of acetic acid bacteria (AAB) in lambic beer production processes, mainly due to their difficult recovery and possibly unknown role. Therefore, a novel aseptic sampling method, spanning both the spatial and temporal distributions of the AAB and their substrates and metabolites, was combined with a highly selective medium and matrix-assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF MS) as a high-throughput dereplication method followed by comparative gene sequencing for their isolation and identification, respectively. The AAB ( species more than species) proliferated during two phases of the lambic beer production process, represented by during a few days in the beginning of the fermentation and from 7 weeks until 24 months of maturation. Competitive exclusion tests combined with comparative genomic analysis of all genomes of strains of both species available disclosed possible reasons for this successive dominance. The spatial analysis revealed that significantly higher concentrations of acetic acid (from ethanol) and acetoin (from lactic acid) were produced at the tops of the casks, due to higher AAB counts and a higher metabolic activity of the AAB species at the air/liquid interface during the first 6 months of lambic beer production. In contrast, no differences in AAB species diversity occurred throughout the casks. Lambic beer is an acidic beer that is the result of a spontaneous fermentation and maturation process. Acidic beers are currently attracting attention worldwide. Part of the acidity of these beers is caused by acetic acid bacteria (AAB). However, due to their difficult recovery, they were never investigated extensively regarding their occurrence, species diversity, and functional role in lambic beer production. In the present study, a framework was developed for their isolation and identification using a novel aseptic sampling method in combination with matrix-assisted laser desorption ionization-time of flight mass spectrometry as a high-throughput dereplication technique followed by accurate molecular identification. The sampling method applied enabled us to take spatial differences into account regarding both enumerations and metabolite production. In this way, it was shown that more AAB were present and more acetic acid was produced at the air/liquid interface during a major part of the lambic beer production process. Also, two different AAB species were encountered, namely, at the beginning and in a later stage of the production process. This developed framework could also be applied for other fermentation processes.
Topics: Acetic Acid; Acetobacter; Beer; Fermentation; Gluconobacter; Microbiota
PubMed: 29352086
DOI: 10.1128/AEM.02846-17 -
MBio Jul 2014Soil microbial diversity represents the largest global reservoir of novel microorganisms and enzymes. In this study, we coupled functional metagenomics and DNA...
Soil microbial diversity represents the largest global reservoir of novel microorganisms and enzymes. In this study, we coupled functional metagenomics and DNA stable-isotope probing (DNA-SIP) using multiple plant-derived carbon substrates and diverse soils to characterize active soil bacterial communities and their glycoside hydrolase genes, which have value for industrial applications. We incubated samples from three disparate Canadian soils (tundra, temperate rainforest, and agricultural) with five native carbon ((12)C) or stable-isotope-labeled ((13)C) carbohydrates (glucose, cellobiose, xylose, arabinose, and cellulose). Indicator species analysis revealed high specificity and fidelity for many uncultured and unclassified bacterial taxa in the heavy DNA for all soils and substrates. Among characterized taxa, Actinomycetales (Salinibacterium), Rhizobiales (Devosia), Rhodospirillales (Telmatospirillum), and Caulobacterales (Phenylobacterium and Asticcacaulis) were bacterial indicator species for the heavy substrates and soils tested. Both Actinomycetales and Caulobacterales (Phenylobacterium) were associated with metabolism of cellulose, and Alphaproteobacteria were associated with the metabolism of arabinose; members of the order Rhizobiales were strongly associated with the metabolism of xylose. Annotated metagenomic data suggested diverse glycoside hydrolase gene representation within the pooled heavy DNA. By screening 2,876 cloned fragments derived from the (13)C-labeled DNA isolated from soils incubated with cellulose, we demonstrate the power of combining DNA-SIP, multiple-displacement amplification (MDA), and functional metagenomics by efficiently isolating multiple clones with activity on carboxymethyl cellulose and fluorogenic proxy substrates for carbohydrate-active enzymes. Importance: The ability to identify genes based on function, instead of sequence homology, allows the discovery of genes that would not be identified through sequence alone. This is arguably the most powerful application of metagenomics for the recovery of novel genes and a natural partner of the stable-isotope-probing approach for targeting active-yet-uncultured microorganisms. We expanded on previous efforts to combine stable-isotope probing and metagenomics, enriching microorganisms from multiple soils that were active in degrading plant-derived carbohydrates, followed by construction of a cellulose-based metagenomic library and recovery of glycoside hydrolases through functional metagenomics. The major advance of our study was the discovery of active-yet-uncultivated soil microorganisms and enrichment of their glycoside hydrolases. We recovered positive cosmid clones in a higher frequency than would be expected with direct metagenomic analysis of soil DNA. This study has generated an invaluable metagenomic resource that future research will exploit for genetic and enzymatic potential.
Topics: Actinomycetales; Caulobacteraceae; Isotope Labeling; Metagenomics; Molecular Sequence Data; Rhodospirillales; Soil Microbiology
PubMed: 25028422
DOI: 10.1128/mBio.01157-14